The Plasminogen–Activator Plasmin System in Physiological and Pathophysiological Angiogenesis

Angiogenesis is a process associated with the migration and proliferation of endothelial cells (EC) to form new blood vessels. It is involved in various physiological and pathophysiological conditions and is controlled by a wide range of proangiogenic and antiangiogenic molecules. The plasminogen activator–plasmin system plays a major role in the extracellular matrix remodeling process necessary for angiogenesis. Urokinase/tissue-type plasminogen activators (uPA/tPA) convert plasminogen into the active enzyme plasmin, which in turn activates matrix metalloproteinases and degrades the extracellular matrix releasing growth factors and proangiogenic molecules such as the vascular endothelial growth factor (VEGF-A). The plasminogen activator inhibitor-1 (PAI-1) is the main inhibitor of uPA and tPA, thereby an inhibitor of pericellular proteolysis and intravascular fibrinolysis, respectively. Paradoxically, PAI-1, which is expressed by EC during angiogenesis, is elevated in several cancers and is found to promote angiogenesis by regulating plasmin-mediated proteolysis and by promoting cellular migration through vitronectin. The urokinase-type plasminogen activator receptor (uPAR) also induces EC cellular migration during angiogenesis via interacting with signaling partners. Understanding the molecular functions of the plasminogen activator plasmin system and targeting angiogenesis via blocking serine proteases or their interactions with other molecules is one of the major therapeutic strategies scientists have been attracted to in controlling tumor growth and other pathological conditions characterized by neovascularization.     

Novel Drugs with High Efficacy against Tumor Angiogenesis

Angiogenesis is involved in physiological and pathological processes in the body. Tumor angiogenesis is a key factor associated with tumor growth, progression, and metastasis. Therefore, there is great interest in developing antiangiogenic strategies. Hypoxia is the basic initiating factor of tumor angiogenesis, which leads to the increase of vascular endothelial growth factor (VEGF), angiopoietin (Ang), hypoxia-inducible factor (HIF-1), etc. in hypoxic cells. The pathways of VEGF and Ang are considered to be critical steps in tumor angiogenesis. A number of antiangiogenic drugs targeting VEGF/VEGFR (VEGF receptor) or ANG/Tie2, or both, are currently being used for cancer treatment, or are still in various stages of clinical development or preclinical evaluation. This article aims to review the mechanisms of angiogenesis and tumor angiogenesis and to focus on new drugs and strategies for the treatment of antiangiogenesis. However, antitumor angiogenic drugs alone may not be sufficient to eradicate tumors. The molecular chaperone heat shock protein 90 (HSP90) is considered a promising molecular target. The VEGFR system and its downstream signaling molecules depend on the function of HSP90. This article also briefly introduces the role of HSP90 in angiogenesis and some HSP90 inhibitors.     

Netrin-1 in Glioblastoma Neovascularization: The New Partner in Crime?

Glioblastoma (GBM) is the most aggressive and common primary tumor of the central nervous system. It is characterized by having an infiltrating growth and by the presence of an excessive and aberrant vasculature. Some of the mechanisms that promote this neovascularization are angiogenesis and the transdifferentiation of tumor cells into endothelial cells or pericytes. In all these processes, the release of extracellular microvesicles by tumor cells plays an important role. Tumor cell-derived extracellular microvesicles contain pro-angiogenic molecules such as VEGF, which promote the formation of blood vessels and the recruitment of pericytes that reinforce these structures. The present study summarizes and discusses recent data from different investigations suggesting that Netrin-1, a highly versatile protein recently postulated as a non-canonical angiogenic ligand, could participate in the promotion of neovascularization processes in GBM. The relevance of determining the angiogenic signaling pathways associated with the interaction of Netrin-1 with its receptors is posed. Furthermore, we speculate that this molecule could form part of the microvesicles that favor abnormal tumor vasculature. Based on the studies presented, this review proposes Netrin-1 as a novel biomarker for GBM progression and vascularization.     

EphrinB2–EphB4 Signaling in Neurooncological Disease

EphrinB2–EphB4 signaling is critical during embryogenesis for cardiovascular formation and neuronal guidance. Intriguingly, critical expression patterns have been discovered in cancer pathologies over the last two decades. Multiple connections to tumor migration, growth, angiogenesis, apoptosis, and metastasis have been identified in vitro and in vivo. However, the molecular signaling pathways are manifold and signaling of the EphB4 receptor or the ephrinB2 ligand is cancer type specific. Here we explore the impact of these signaling pathways in neurooncological disease, including glioma, brain metastasis, and spinal bone metastasis. We identify potential downstream pathways that mediate cancer suppression or progression and seek to understand it´s role in antiangiogenic therapy resistance in glioma. Despite the Janus-faced functions of ephrinB2–EphB4 signaling in cancer Eph signaling remains a promising clinical target.     

Nanomaterials for Antiangiogenic Therapies for Cancer: A Promising Tool for Personalized Medicine

Angiogenesis is one of the hallmarks of cancer. Several studies have shown that vascular endothelium growth factor (VEGF) plays a leading role in angiogenesis progression. Antiangiogenic medication has gained substantial recognition and is commonly administered in many forms of human cancer, leading to a rising interest in cancer therapy. However, this treatment method can lead to a deteriorating outcome of resistance, invasion, distant metastasis, and overall survival relative to its cytotoxicity. Furthermore, there are significant obstacles in tracking the efficacy of antiangiogenic treatments by incorporating positive biomarkers into clinical settings. These shortcomings underline the essential need to identify additional angiogenic inhibitors that target numerous angiogenic factors or to develop a new method for drug delivery of current inhibitors. The great benefits of nanoparticles are their potential, based on their specific properties, to be effective mechanisms that concentrate on the biological system and control various important functions. Among various therapeutic approaches, nanotechnology has emerged as a new strategy for treating different cancer types. This article attempts to demonstrate the huge potential for targeted nanoparticles and their molecular imaging applications. Notably, several nanoparticles have been developed and engineered to demonstrate antiangiogenic features. This nanomedicine could effectively treat a number of cancers using antiangiogenic therapies as an alternative approach. We also discuss the latest antiangiogenic and nanotherapeutic strategies and highlight tumor vessels and their microenvironments.     

Nitric Oxide Modulates Metabolic Processes in the Tumor Immune Microenvironment

The metabolic requirements and functions of cancer and normal tissues are vastly different. Due to the rapid growth of cancer cells in the tumor microenvironment, distorted vasculature is commonly observed, which creates harsh environments that require rigorous and constantly evolving cellular adaption. A common hallmark of aggressive and therapeutically resistant tumors is hypoxia and hypoxia-induced stress markers. However, recent studies have identified alterations in a wide spectrum of metabolic pathways that dictate tumor behavior and response to therapy. Accordingly, it is becoming clear that metabolic processes are not uniform throughout the tumor microenvironment. Metabolic processes differ and are cell type specific where various factors promote metabolic heterogeneity within the tumor microenvironment. Furthermore, within the tumor, these metabolically distinct cell types can organize to form cellular neighborhoods that serve to establish a pro-tumor milieu in which distant and spatially distinct cellular neighborhoods can communicate via signaling metabolites from stroma, immune and tumor cells. In this review, we will discuss how biochemical interactions of various metabolic pathways influence cancer and immune microenvironments, as well as associated mechanisms that lead to good or poor clinical outcomes.     

Escaping Antiangiogenic Therapy: Strategies Employed by Cancer Cells

Tumor angiogenesis is widely recognized as one of the “hallmarks of cancer”. Consequently, during the last decades the development and testing of commercial angiogenic inhibitors has been a central focus for both basic and clinical cancer research. While antiangiogenic drugs are now incorporated into standard clinical practice, as with all cancer therapies, tumors can eventually become resistant by employing a variety of strategies to receive nutrients and oxygen in the event of therapeutic assault. Herein, we concentrate and review in detail three of the principal mechanisms of antiangiogenic therapy escape: (1) upregulation of compensatory/alternative pathways for angiogenesis; (2) vasculogenic mimicry; and (3) vessel co-option. We suggest that an understanding of how a cancer cell adapts to antiangiogenic therapy may also parallel the mechanisms employed in the bourgeoning tumor and isolated metastatic cells delivering responsible for residual disease. Finally, we speculate on strategies to adapt antiangiogenic therapy for future clinical uses.     

AS1411 Nucleolin-Specific Binding Aptamers Reduce Pathological Angiogenesis through Inhibition of Nucleolin Phosphorylation

Proliferative retinopathies produces an irreversible type of blindness affecting working age and pediatric population of industrialized countries. Despite the good results of anti-VEGF therapy, intraocular and systemic complications are often associated after its intravitreal use, hence novel therapeutic approaches are needed. The aim of the present study is to test the effect of the AS1411, an antiangiogenic nucleolin-binding aptamer, using in vivo, ex vivo and in vitro models of angiogenesis and propose a mechanistic insight. Our results showed that AS1411 significantly inhibited retinal neovascularization in the oxygen induced retinopathy (OIR) in vivo model, as well as inhibited branch formation in the rat aortic ex vivo assay, and, significantly reduced proliferation, cell migration and tube formation in the HUVEC in vitro model. Importantly, phosphorylated NCL protein was significantly abolished in HUVEC in the presence of AS1411 without affecting NFκB phosphorylation and -21 and 221-angiomiRs, suggesting that the antiangiogenic properties of this molecule are partially mediated by a down regulation in NCL phosphorylation. In sum, this new research further supports the NCL role in the molecular etiology of pathological angiogenesis and identifies AS1411 as a novel anti-angiogenic treatment.     

Key Role of Astrocytes in Postnatal Brain and Retinal Angiogenesis

Angiogenesis is a key process in various physiological and pathological conditions in the nervous system and in the retina during postnatal life. Although an increasing number of studies have addressed the role of endothelial cells in this event, the astrocytes contribution in angiogenesis has received less attention. This review is focused on the role of astrocytes as a scaffold and in the stabilization of the new blood vessels, through different molecules release, which can modulate the angiogenesis process in the brain and in the retina. Further, differences in the astrocytes phenotype are addressed in glioblastoma, one of the most devastating types of brain cancer, in order to provide potential targets involved in the cross signaling between endothelial cells, astrocytes and glioma cells, that mediate tumor progression and pathological angiogenesis. Given the relevance of astrocytes in angiogenesis in physiological and pathological conditions, future studies are required to better understand the interrelation between endothelial and astrocyte signaling pathways during this process.     

Hierarchical mechanochemical switches in angiostatin

We wish to propose a novel mechanism by which the triggering of a biochemical signal can be controlled by the hierarchical coupling between a protein redox equilibrium and an external mechanical force. We have characterized this mechanochemical mechanism in angiostatin, and we have evidence that it can switch the access to partially unfolded structures of this protein. We have identified a metastable intermediate that is specifically accessible under thioredoxin-rich reducing conditions, like those met by angiostatin on the surface of a tumor cell. The structure of the same intermediate accounts for the unexplained antiangiogenic activity of angiostatin. These findings demonstrate a new link between redox biology and mechanically regulated processes.     

Systematic Review on Tumor Microenvironment in Glial Neoplasm: From Understanding Pathogenesis to Future Therapeutic Perspectives

Despite the multidisciplinary management in the treatment of glioblastomas, the average survival of GBM patients is still 15 months. In recent years, molecular biomarkers have gained more and more importance both in the diagnosis and therapy of glial tumors. At the same time, it has become clear that non neoplastic cells, which constitute about 30% of glioma mass, dramatically influence tumor growth, spread, and recurrence. This is the main reason why, in recent years, scientific research has been focused on understanding the function and the composition of tumor microenvironment and its role in gliomagenesis and recurrence. The aim of this review is to summarize the most recent discovery about resident microglia, tumor-associated macrophages, lymphocytes, and the role of extracellular vesicles and their bijective interaction with glioma cells. Moreover, we reported the most recent updates about new therapeutic strategies targeting immune system receptors and soluble factors. Understanding how glioma cells interact with non-neoplastic cells in tumor microenvironment is an essential step to comprehend mechanisms at the base of disease progression and to find new therapeutic strategies for GBM patients. However, no significant results have yet been obtained in studies targeting single molecules/pathways; considering the complex microenvironment, it is likely that only by using multiple therapeutic agents acting on multiple molecular targets can significant results be achieved.     

LPPR5 Expression in Glioma Affects Growth,Vascular Architecture,and Sunitinib Resistance

Despite intensive research, glioblastoma remains almost invariably fatal. Various promising drugs targeting specific aspects of glioma biology, in addition to or as an alternative to antiproliferative chemotherapy, were not successful in larger clinical trials. Further insights into the biology of glioma and the mechanisms behind the evasive-adaptive response to targeted therapies is needed to help identify new therapeutic targets, prognostics, or predictive biomarkers. As a modulator of the canonically oncogenic Rho-GTPase pathway, Lipid phosphate phosphatase-related protein type 5 (LPPR5) is pivotal in influencing growth, angiogenesis, and therapeutic resistance. We used a GL261 murine orthotopic allograft glioma model to quantify the tumor growth and to obtain tissue for histological and molecular analysis. Epicortical intravital epi-illumination fluorescence video microscopy of the tumor cell spheroids was used to characterize the neovascular architecture and hemodynamics. GL261-glioma growth was delayed and decelerated after LPPR5 overexpression (LPPR5^OE). We observed increased tumor cell apoptosis and decreased expression and secretion of vascular endothelial growth factor A in LPPR5^OE glioma. Hence, an altered micro-angioarchitecture consisting of dysfunctional small blood vessels was discovered in the LPPR5^OE tumors. Sunitinib therapy eliminated these vessels but had no effect on tumor growth or apoptosis. In general, LPPR5 overexpression generated a more benign, proapoptotic glioma phenotype with delayed growth and a dysfunctional vascular architecture.     

The “Angiogenic Switch” and Functional Resources in Cyclic Sports Athletes

Regular physical activity in cyclic sports can influence the so-called “angiogenic switch”, which is considered as an imbalance between proangiogenic and anti-angiogenic molecules. Disruption of the synthesis of angiogenic molecules can be caused by local changes in tissues under the influence of excessive physical exertion and its consequences, such as chronic oxidative stress and associated hypoxia, metabolic acidosis, sports injuries, etc. A review of publications on signaling pathways that activate and inhibit angiogenesis in skeletal muscles, myocardium, lung, and nervous tissue under the influence of intense physical activity in cyclic sports. Materials: We searched PubMed, SCOPUS, Web of Science, Google Scholar, Clinical keys, and e-LIBRARY databases for full-text articles published from 2000 to 2020, using keywords and their combinations. Results: An important aspect of adaptation to training loads in cyclic sports is an increase in the number of capillaries in muscle fibers, which improves the metabolism of skeletal muscles and myocardium, as well as nervous and lung tissue. Recent studies have shown that myocardial endothelial cells not only respond to hemodynamic forces and paracrine signals from neighboring cells, but also take an active part in heart remodeling processes, stimulating the growth and contractility of cardiomyocytes or the production of extracellular matrix proteins in myofibroblasts. As myocardial vascularization plays a central role in the transition from adaptive heart hypertrophy to heart failure, further study of the signaling mechanisms involved in the regulation of angiogenesis in the myocardium is important in sports practice. The study of the “angiogenic switch” problem in the cerebrovascular and cardiovascular systems allows us to claim that the formation of new vessels is mediated by a complex interaction of all growth factors. Although the lungs are one of the limiting systems of the body in cyclic sports, their response to high-intensity loads and other environmental stresses is often overlooked. Airway epithelial cells are the predominant source of several growth factors throughout lung organogenesis and appear to be critical for normal alveolarization, rapid alveolar proliferation, and normal vascular development. There are many controversial questions about the role of growth factors in the physiology and pathology of the lungs. The presented review has demonstrated that when doing sports, it is necessary to give a careful consideration to the possible positive and negative effects of growth factors on muscles, myocardium, lung tissue, and brain. Primarily, the “angiogenic switch” is important in aerobic sports (long distance running). Conclusions: Angiogenesis is a physiological process of the formation of new blood capillaries, which play an important role in the functioning of skeletal muscles, myocardium, lung, and nervous tissue in athletes. Violation of the “angiogenic switch” as a balance between proangiogenic and anti-angiogenic molecules can lead to a decrease in the functional resources of the nervous, musculoskeletal, cardiovascular, and respiratory systems in athletes and, as a consequence, to a decrease in sports performance.     

Toward Angiogenesis Inhibitors Based on the Conjugation of Organometallic Platinum(II) Complexes to RGD Peptides

A novel conjugate between a cyclometalated platinum(II) complex with dual antiangiogenic and antitumor activity and a cyclic peptide containing the RGD sequence (‐Arg‐Gly‐Asp‐) has been synthesized by combining solid‐ and solution‐phase methodologies. Although peptide conjugation rendered a non‐cytotoxic compound in all tested tumor cell lines (± α_Vβ₃ and α_Vβ₅ integrin receptors), the antiangiogenic activity of the Pt‐c(RGDfK) conjugate in human umbilical vein endothelial cells at sub‐cytotoxic concentrations opens the way to the design of a novel class of angiogenesis inhibitors through conjugation of metallodrugs with high antiangiogenic activity to cyclic RGD‐containing peptides or peptidomimetic analogues.     

Differential Angiogenic Potential of 3-Dimension Spheroid of HNSCC Cells in Mouse Xenograft

The experimental animal model is still essential in the development of new anticancer drugs. We characterized mouse tumors derived from two-dimensional (2D) monolayer cells or three-dimensional (3D) spheroids to establish an in vivo model with highly standardized conditions. Primary cancer-associated fibroblasts (CAFs) were cultured from head and neck squamous cell carcinoma (HNSCC) tumor tissues and co-injected with monolayer cancer cells or spheroids into the oral mucosa of mice. Mice tumor blood vessels were stained, followed by tissue clearing and 3D Lightsheet fluorescent imaging. We compared the effect of exosomes secreted from 2D or 3D culture conditions on the angiogenesis-related genes in HNSCC cells. Our results showed that both the cells and spheroids co-injected with primary CAFs formed tumors. Interestingly, vasculature was abundantly distributed inside the spheroid-derived but not the monolayer-derived mice tumors. In addition, cisplatin injection more significantly decreased spheroid-derived but not monolayer-derived tumor size in mice. Additionally, exosomes isolated from co-culture media of FaDu spheroid and CAF upregulated angiogenesis-related genes in HNSCC cells as compared to exosomes from FaDu cell and CAF co-culture media under in vitro conditions. The mouse tumor xenograft model derived from 3D spheroids of HNSCC cells with primary CAFs is expected to produce reliable chemotherapy drug screening results given the robust angiogenesis and lack of necrosis inside tumor tissues.     

Intratumoral Canine Distemper Virus Infection Inhibits Tumor Growth by Modulation of the Tumor Microenvironment in a Murine Xenograft Model of Canine Histiocytic Sarcoma

Histiocytic sarcomas refer to highly aggressive tumors with a poor prognosis that respond poorly to conventional treatment approaches. Oncolytic viruses, which have gained significant traction as a cancer therapy in recent decades, represent a promising option for treating histiocytic sarcomas through their replication and/or by modulating the tumor microenvironment. The live attenuated canine distemper virus (CDV) vaccine strain Onderstepoort represents an attractive candidate for oncolytic viral therapy. In the present study, oncolytic virotherapy with CDV was used to investigate the impact of this virus infection on tumor cell growth through direct oncolytic effects or by virus-mediated modulation of the tumor microenvironment with special emphasis on angiogenesis, expression of selected MMPs and TIMP-1 and tumor-associated macrophages in a murine xenograft model of canine histiocytic sarcoma. Treatment of mice with xenotransplanted canine histiocytic sarcomas using CDV induced overt retardation in tumor progression accompanied by necrosis of neoplastic cells, increased numbers of intratumoral macrophages, reduced angiogenesis and modulation of the expression of MMPs and TIMP-1. The present data suggest that CDV inhibits tumor growth in a multifactorial way, including direct cell lysis and reduction of angiogenesis and modulation of MMPs and their inhibitor TIMP-1, providing further support for the concept of its role in oncolytic therapies.     

Tumor-Associated Macrophages (TAMs) in Colorectal Cancer (CRC): From Mechanism to Therapy and Prognosis

Colorectal cancer (CRC) is a malignant tumor in the digestive system whose incidence and mortality is high-ranking among tumors worldwide. The initiation and progression of CRC is a complex process involving genetic alterations in cancer cells and multiple factors from the surrounding tumor cell microenvironment. As accumulating evidence has shown, tumor-associated macrophages (TAMs)—as abundant and active infiltrated inflammatory cells in the tumor microenvironment (TME)—play a crucial role in CRC. This review focuses on the different mechanisms of TAM in CRC, including switching of phenotypical subtypes; promoting tumor proliferation, invasion, and migration; facilitating angiogenesis; mediating immunosuppression; regulating metabolism; and interacting with the microbiota. Although controversy remains in clinical evidence regarding the role of TAMs in CRC, clarifying their significance in therapy and the prognosis of CRC may shed new light on the optimization of TAM-centered approaches in clinical care.     

In Vitro Antimetastatic Effect of Phosphatidylinositol 3-Kinase Inhibitor ZSTK474 on Prostate Cancer PC3 Cells

Tumor metastasis is the main cause of lethality of prostate cancer, because conventional therapies like surgery and hormone treatment rarely work at this stage. Tumor cell migration, invasion and adhesion are necessary processes for metastasis. By providing nutrition and an escape route from the primary site, angiogenesis is also required for tumor metastasis. Phosphatidylinositol 3-kinases (PI3Ks) are well known to play important roles in tumorigenesis as well as metastasis. ZSTK474 is a specific PI3K inhibitor developed for solid tumor therapy. In the present report, antimetastatic activities of ZSTK474 were investigated in vitro by determining the effects on the main metastatic processes. ZSTK474 exhibited inhibitory effects on migration, invasion and adhesive ability of prostate cancer PC3 cells. Furthermore, ZSTK474 inhibited phosphorylation of Akt substrate-Girdin, and the secretion of matrix metalloproteinase (MMP), both of which were reported to be closely involved in migration and invasion. On the other hand, ZSTK474 inhibited the expression of HIF-1α and the secretion of vascular endothelial growth factor (VEGF), suggesting its potential antiangiogenic activity on PC3 cells. Moreover, we demonstrated the antiangiogenesis by determining the effect of ZSTK474-reduced VEGF on tube formation of human umbilical vein endothelial cells (HUVECs). In conclusion, ZSTK474 was demonstrated to have potential in vitro antimetastatic effects on PC3 cells via dual mechanisms: inhibition of metastatic processes including cell migration, invasion and adhesion, and antiangiogenesis via blockade of VEGF secretion.     

Overcoming Radiation Resistance in Gliomas by Targeting Metabolism and DNA Repair Pathways

Gliomas represent a wide spectrum of brain tumors characterized by their high invasiveness, resistance to chemoradiotherapy, and both intratumoral and intertumoral heterogeneity. Recent advances in transomics studies revealed that enormous abnormalities exist in different biological layers of glioma cells, which include genetic/epigenetic alterations, RNA expressions, protein expression/modifications, and metabolic pathways, which provide opportunities for development of novel targeted therapeutic agents for gliomas. Metabolic reprogramming is one of the hallmarks of cancer cells, as well as one of the oldest fields in cancer biology research. Altered cancer cell metabolism not only provides energy and metabolites to support tumor growth, but also mediates the resistance of tumor cells to antitumor therapies. The interactions between cancer metabolism and DNA repair pathways, and the enhancement of radiotherapy sensitivity and assessment of radiation response by modulation of glioma metabolism are discussed herein.     

Fatty Acids,CD36, Thrombospondin-1, and CD47 in Glioblastoma: Together and/or Separately?

Glioblastoma (GBM) is one of the most aggressive tumors of the central nervous system, characterized by a wide range of inter- and intratumor heterogeneity. Accumulation of fatty acids (FA) metabolites was associated with a low survival rate in high-grade glioma patients. The diversity of brain lipids, especially polyunsaturated fatty acids (PUFAs), is greater than in all other organs and several classes of proteins, such as FA transport proteins (FATPs), and FA translocases are considered principal candidates for PUFAs transport through BBB and delivery of PUFAs to brain cells. Among these, the CD36 FA translocase promotes long-chain FA uptake as well as oxidated lipoproteins. Moreover, CD36 binds and recognizes thrombospondin-1 (TSP-1), an extracellular matrix protein that was shown to play a multifaceted role in cancer as part of the tumor microenvironment. Effects on tumor cells are mediated by TSP-1 through the interaction with CD36 as well as CD47, a member of the immunoglobulin superfamily. TSP-1/CD47 interactions have an important role in the modulation of glioma cell invasion and angiogenesis in GBM. Separately, FA, the two membrane receptors CD36, CD47, and their joint ligand TSP-1 all play a part in GBM pathogenesis. The last research has put in light their interconnection/interrelationship in order to exert a cumulative effect in the modulation of the GBM molecular network.